In order to address the relevance of phospholipase C (PLC)-gamma activity in mitogen-initiated proliferative signals, we overexpressed PLC-gamma in NIH/3T3 fibroblasts by transfecting the bovine cDNA under the transcriptional control of the murine leukemia virus long terminal repeats. The amount of PLC-gamma recovered by antiphosphotyrosine immunoprecipitation was increased in overexpressor cells after treatment with platelet-derived growth factor (PDGF) and, to a lesser extent. with basic fibroblast growth factor (FGF). This correlated with increased phosphoinositol release induced by PDGF and basic FGF in the PLC-gamma overexpressing cells. These findings support the concept that PLC-gamma is rapidly activated in fibroblasts following stimulation with PDGF and basic FGF and suggest that phosphatidyl inositol breakdown is one limiting step in transduction of the proliferative signal initiated by these two growth factors. The role of protein kinase C (PKC)-gamma in tumorigenesis was analyzed by overexpressing PKC-gamma in NIH/3T3 fibroblasts. Different degrees of overexpression were achieved, ranging from twofold to 30-fold as determined by immunoblot analysis. Cells containing 20-fold more PKC than normal exhibited the transformed phenotype as determined by cell morphology. saturation density. and growth in soft agar. Interestingly, the malignant properties of these cells, as well as of the human overexpressor ones, were drastically reduced following chronic treatment with phorbol esters. In addition, saturation density and morphology were more similar to normal cells when grown in serum-free conditions. NIH/3T3 fibroblasts with two- to 10-fold more PKC than normal exhibited an enhanced mitogenic response to specific activators of PKC. These results indicate that PKC exerts its transforming properties by amplifying the cell response to external mitogenic stimuli.